The invention relates generally to a method for modulating intraocular pressure (IOP) in an eye of a human or non-human mammalian subject susceptible to or having elevated IOP, and more particularly to a method for modulating the IOP by treating the eye with an inhibitor of integrin-linked kinase.
When aqueous humor cannot drain normally from the anterior chamber of an eye, an animal can develop one of a family of ophthalmologic disorders. These disorders are characterized by above normal IOP and gradual neuropathy caused in some manner by increased pressure on the optic nerve. Pressure increase begins in the anterior chamber and extends to the other parts of the eye, including the posterior chamber. Under the force of the IOP, the posterior chamber compresses and destroys nerve fibers and blood vessels of the optic nerve. Such disorders can lead to gradual visual impairment and are collectively referred to as glaucoma. In a healthy mammalian eye, the aqueous humor is under resistance from the trabecular meshwork structures, generating a normal physiological IOP in a range from about 12-20 mmHg. The resistance directs and regulates outflow of aqueous humor from the eye.
Integrins are believed to help regulate IOP. They are found on virtually all human cells and transmit signals bi-directionally across a cell membrane. As cell surface receptors, integrins participate in a diverse array of biological functions including cellular development, cellular/tissue repair, angiogenesis, inflammation and hemostasis.
Integrin structure and function are known in the art. See Alpin A., et al., “Signal transduction and signal modulation by cell adhesion receptors: the role of integrins, cadherins, immunoglobulin-cell adhesion molecules, and selectins,” Pharmacological Reviews 50: 197-263 (1998); and Brakebusch C. & Fässler R., “The integrin-actin connection, an eternal love affair,” EMBO Journal 22:2324-2333 (2003), each of which is incorporated herein by reference as if set forth in its entirety. At least eighteen isoforms of the α-subunit and eight isoforms of the β-subunit combine to form more than twenty integrin heterodimers having α- and β-subunits. Human trabecular meshwork cells contain the following integrin subunits: α1, α3, α4, α5, α6, αv, β1, β3, β4 and α5. Zhou L., et al., “Expression of integrin receptors in the human trabecular meshwork,” Current Eye Research 19:395-402 (1999). Each integrin heterodimer has an extracellular domain, a transmembrane domain and a cytoplasmic tail.
The extracellular domain participates in cellular adhesion by binding to an Arg-Gly-Asp (RGD) amino acid sequence in a ligand. Known integrin ligands are ECM proteins including, but not limited to, vitronectin, fibronectin, type I and IV collagen and vascular cell adhesion molecule.
The cytoplasmic tail interacts with more than twenty intracellular constituents, linking the integrin to the actin cytoskeleton and forming focal adhesions, focal complexes and fibrillar adhesions. Signaling pathways also linked to the cytoplasmic tail of integrins are MAP kinase, FAK, JAK-STAT, JNK, inositol lipid pathway and Rho family of GTPases (including Rac and Cdc42).
Fibronectin interacts with cell surface receptors via two of its domains: (1) a central cell binding domain (CBD) and (2) a carboxy-terminal heparin-binding domain (Hep II). The CBD contains an RGD amino acid sequence that binds to an integrin; whereas, the Hep II domain contains three sequences that bind to cell surface receptors. Hep II has an amino acid sequence that binds to heparin-sulfate groups on a syndecan cell-surface receptor as well as an amino acid sequence (IDAPS) that binds to integrins. A third site called IIICS (type III) connecting sequence also contains a cell binding domain. This domain contains binding sites for integrins (LDV) as well as for cell surface proteoglycans.
The CBD and the Hep II domain together bring the integrin and the syndecan into proximity to mediate an intracellular signaling event that alters a cell's cytoskeleton. Cell signaling, mediated by the binding of fibronectin to integrins, involves integrin-linked kinase (ILK), a 59 kDa intracellular, cell-signaling enzyme. ILK interacts with the cytoplasmic tail of β1- or β3-subunits of integrin heterodimers. ILK also associates with other adaptor and signaling proteins such as PINCH, CH-ILKBP, affixin and paxillin, and with catalytic proteins such as ILKAP, PKB/Akt and PDK-1. See Wu C. & Dedhar S., “Integrin-linked kinase (ILK) and its interactors: a new paradigm for the coupling of extracellular matrix to actin cytoskeleton and signaling complexes,” J. Cell. Biology 155:505-510 (2001), incorporated herein by reference as if set forth in its entirety. See also, Khyrul W., “The integrin-linked kinase regulates cell morphology and motility in a Rho-associated kinase- dependent manner,” J. B. C. 279:54131-54139 (2004), incorporated herein by reference as if set forth in its entirety.
ILK phosphorylates serine/threonine residues on other cell signaling molecules. ILK itself, however, must first be phosphorylated by phosphotidylinositol-3-kinase (PI3K) or by auto-phosphorylation. Conversely, ILK is negatively regulated by phosphatases, including PTEN and ILKAP. The cell-signaling molecules activated by ILK regulate cell survival, cell adhesion and ECM modification. ILK signaling also affects the regulation of cell migration, cell motility and contractility and is involved in suppressing apoptosis and in advancing the cell cycle.
Integrins, fibronectin and ILK interact to form focal adhesions, which are macromolecular complexes found where cells adhere to the extracellular matrix. Focal adhesions are linked to actin stress fibers and serve as signaling complexes for triggering intracellular cascades. Cell-to-ECM interactions with integrins, fibronectin and ILK are implicated in a variety of pathophysiological conditions, including glaucoma.
Glaucoma can be classified into two broad classes—open-angle and closed-angle glaucoma, each of which is subclassified into primary and secondary forms. In primary open-angle glaucoma (also known as POAG), the trabecular meshwork appears to not function properly. Aqueous humor outflow from the eye is restricted. Consequently, the aqueous humor builds up in the anterior chamber, increasing IOP because it cannot flow through the trabecular meshwork. The cause of this reduced outflow is not known. In secondary open-angle glaucoma, intraocular inflammation or the use of certain treatments such as steroids can increase IOP.
Current treatments for glaucoma include pharmacological and surgical therapies, either alone or in combination. All treatments can have significant side effects. Pharmacological agents, most commonly administered as eye drops, can be used alone or in combination to decrease aqueous humor production or to improve aqueous humor outflow from the eye. β-adrenergic blockers such as timolol, levobunolol and betaxolol decrease aqueous humor production. Side effects of β-adrenergic blockers can include cardiac failure, heart block and bronchospasm. Cholinergic agonists such as pilocarpine, carbachol, and phospholine iodide improve outflow facility from the trabecular meshwork. Side effects of cholinergic agonists can include miosis, brow ache and decreased vision. Carbonic anhydrase inhibitors such as acetazolamide, dorzolamide and brinzolamide decrease aqueous humor production. Side effects of carbonic anhydrase inhibitors can include gastrointestinal upset, malaise, renal stones and aplastic anemia. Non-selective α-agonists such as epinephrine and dipivefrin decrease aqueous humor production and increase trabecular outflow facility. Side effects of non-selective α-agonists can include pupil dilation, macular edema and tachycardia. Selective α-agonists such as apraclonidine and brimonidine decrease aqueous humor production and increase outflow through the uveoscleral pathway (an alternative, but smaller, fluid exit pathway to the trabecular meshwork). Side effects of selective α-agonists can include contact allergy and hypotension. Prostaglandin agonists such as latanoprost, travoprost and bimatoprost improve uveoscleral outflow. Side effects of prostaglandins can include iris color change, lash growth and trichiasis. Hyperosmotics such as glycerin (po) and mannitol (iv) establish a concentration gradient that draws excess aqueous humor from the eye. Side effects of using hyperosmotics can include diuresis, cardiovascular overload, renal insufficiency, and stroke, so their use is limited to emergency situations.
When pharmacological agents are unsuccessful in open-angle glaucoma or when a subject presents with closed-angle glaucoma, invasive surgery is indicated. In argon laser trabeculoplasty (ALT), a laser beam is directed at the trabecular meshwork that increases aqueous humor drainage through a mechanism that is not well understood. In laser cyclophotocoagulation, thermal energy applied to the ciliary body destroys the tissue, thereby reducing aqueous humor production. Trabeculectomy establishes a flow route that bypasses the trabecular meshwork so that aqueous humor drains from the anterior chamber just beneath the conjunctiva, the outermost covering of the eye, on the surface of the eye where it is gradually absorbed by blood vessels or diffuses through the conjunctiva. Iridotomy, generally used for closed-angle glaucoma, employs a laser to make an incision in a peripheral area of the iris of the eye to establish a direct aqueous humor flow route between the anterior chamber and the posterior chamber. Iridectomy is similar to iridotomy, but does not employ a laser. In iridotomy, a small section of peripheral iris is surgically excised.
Glaucoma is an increasingly important public health concern, especially in view of the aging of the population. Present treatments do not always adequately control glaucoma, especially steroid-induced glaucoma. Thus, there is a strong need to develop additional methods to prevent and treat the various forms of glaucoma.